Chapter 4 study guide Flashcards
The function of a protein depends on the detailed structure of
its 3-dimensional
shape
A protein or polypeptide is a long chain of amino acids linked together by
covalent peptide bonds
- The 20 different amino acids differ from one another in the chemical structure of their
side chains
Protein folding is influenced by three types of…
non covalent bonds: hydrogen bonds, electrostatic interactions, and van der Waals
Protein folding is also influenced by
hydophobic interactions (nonpolar side chains cluster together)
Polar side chains tend to be found
on the outside surface
The final folded structure or conformation of a polypeptide chain is usually determined
by its
lowest free energy state
Proteins can be unfolded or denatured with solvents that
disrupt non-covalent
interactions
When proteins fold incorrectly, they often form
insoluble aggregates
Many neurodegenerative disorders are associated
with the formation of
protein aggregates
Function of Protein aggregates
kill cells and tissues
unusual type of neurodegenerative disease that is caused by infectious proteins that stimulate protein misfolding
Prion diseases
Relatively easy now to determine a polypeptide sequence by sequencing
its gene
The alpha helix and beta sheet are formed by
hydrogen bonds between N-H and C=O groups in the polypeptide backbone
-the alpha helix (like a spiral staircase) forms a hydrogen bond between every __ amino acid, forming a regular helix with a turn every __ amino acids.
4th, 3.6
alpha helices that span a membrane lipid bilayer typically have __ hydrophobic
amino acids that can contact the hydrophobic lipid tails
20
some alpha helices can wrap around each other to form
coiled coils
The hydrophobic amino acids on one side of an alpha-helix at positions ___ interact with the hydrophobic amino acids on one side of another alpha helix, whereas the hydrophilic amino acids can be exposed to the aqueous environment
a and d
When the polypeptide strands run in the same direction (N to C terminal), they are called
parallel
Most proteins contain small stretches of __ amino acids known as __ that can fold __ into a stable, compact structure.
100-250, protein
domains, independently
The full three-dimensional structure of a polypeptide chain, with its multiple secondary
structures and domains, is considered its
tertiary structure
If a protein molecule contains more than one polypeptide structure, this is known as its
quarternary structure
A polypeptide that is n amino acids long would have __ potential amino acid sequences
20n
Small changes in a polypeptide sequence (mutations) can disrupt protein structure and
function: what’s an example
Sickle cell anemia
Protein families share…
both sequence and structural similarities, but differ in functions
__ are a family of proteins that can cleave peptide bonds in a polypeptide
chain.
serine proteases
A binding site is any region on a protein surface that interacts with another molecule through
non-covalent bonds
In larger protein complexes, each polypeptide chain
is referred to as a
subunit
Most proteins are
globular
fold up into compact shapes like a ball with an irregular surface
globular proteins
Collagen is composed of __ polypeptides that form a __ helix
3, triple
The collagen molecules then bind end-to-end and side-to-side to form
Collagen fibrils
The interactions between the collagen molecules are stabilized by
covalent bonds outside the cell
There are many different kinds of covalent cross-links, but __ between the __ groups of __ are among the most common
disulfide bonds (S-S), SH of cytsesines
These bonds are formed in the lumen of the ER
covalent cross links, S-S bonds
Disulfide bonds are found in many
extracellular proteins, where they stabilize protein conformation
Binding sites depend on the
3D organization of the polypeptide
an example of proteins that bind their ligands with high affinity and specificity
Antibodies (IgG), ligands are antigens
IgGs contain how many large and light chains
2 large and 2 light
IgGs are held together by
S-S bonds
Most of the polypeptide sequence is conserved between what region
constant regions
the antigen-binding site is formed by
small variable loops in heavy and light chains
Changing the length and sequence of the variable loops will alter
binding specificities for different antigens without affecting the structure of the rest of the antibody molecule
breaks down polysaccharide chains in the cell walls of bacteria
Lysozyme
Enzymes make and break
covalent bonds
Many drugs work by blocking the activity of an
enzyme
How are proteins controlled…
- Regulate expression of the gene encoding the protein.
- Target a protein to a specific cellular compartment
- Alter protein degradation
- Turn protein activities on and off by ligand bindings
second sites where regulatory molecules can bind and alter protein conformation to either inactivate or stimulate a protein (or enzyme).
Allosteric regulation
Examples of allosteric regulation
Feedback inhibition (neg and pos)
major mechanism used by cells to control protein activity.
phosphorylation
Phosphate groups contain
2 negative charges and their addition to serine, threonine, or tyrosine side chains alter protein conformation
Phosphate addition is catalyzed by a
kinase
Phosphates are removed from amino acid side chains by a
protein phosphatase
Phosphorylation may either
stimulate or inhibit protein activity
molecular switches that are regulated by the cyclic gain
and loss of a phosphate group
GTP binding sites
Use the energy of ATP binding, hydrolysis, and release to drive the conformational changes needed to move in one direction along a track are done by what proteins
Motor proteins
Ex of motor proteins
myosin moving on an actin filament, kinesin moving on microtubules, DNA polymerase moving on a DNA strand.
Proteins often form large machines with
10 or more subunits
contain binding sites that are recognized by multiple proteins
scaffold proteins
Binding of several proteins to a scaffold enables formation of
protein complex
__ modifications of amino acids after __ also control the location and
assembly of protein machines
covalent, translation
Addition of the __ drives proteins to cell membranes
palmitate to cysteines
The set of covalent modifications that a protein contains is an important way of regulating the protein’s function and constitutes a
regulatory protein code
To isolate a protein from a particular cell type, one must break open the cell (poke holes in the cell membrane) to make a
cell homogenate or extract that contains all the cell
organelles
There are several methods commonly used in cell fractionation and centrifugation
a) sonication (high frequency sound)
b) mild detergent treatment
c) forcing cells through small holes under high pressure
d) mechanical shear
Once you have your cell extract, you can separate it into crude fractions by
differential
centrifugation
Less dense components of centrifuge settle at
the bottom more quickly
Centrifugation: contains whole cells, partially disrupted cells (cell ghosts), and nuclei
low speed pellet
contains the larger organelles (mitochondria, lysosomes,
peroxisomes)
medium speed pellet
contains the larger organelles (mitochondria, lysosomes,
peroxisomes)
high speed pellet
contains large cytoplasmic protein complexes (ribosomes,
virus particles, large macromolecules)
very high speed pellet
Typically used to separate larger protein complexes based on
the rates that they move through a shallow gradient of sucrose.
Velocity sedimentation
Larger, denser complexes will move __ than smaller, less dense complexes
faster
used to separate components based on their buoyant
density, independent of size and shape.
Equilibrium centrifugation
often used to isolate nucleic acids (DNA, RNA)
Equilibrium centrifugation
separates proteins according to their size
Gel filtration chromatography
separate the crude protein mixture on the basis of size, charge, and/or its ability to bind to a particular chemical group
Column chromatography
contain a matrix that is covalently bound to a molecule that interacts
specifically with the protein of interest
Affinity columns
This molecule may resemble an enzyme’s substrate, or it might be a specific antibody that recognizes an antigen on the protein surface
Affinity columns
the proteins are treated with a
negatively charged detergent, sodium dodecyl sulfate (SDS), which denatures the
proteins, and a reducing agent, which breaks disulfide bonds
SDS polyacrylamide gel electrophoresis (SDS-PAGE)
the gel contains a pH gradient, and the proteins are NOT treated with SDS. Proteins will migrate in the pH gradient until they reach their
individual isoelectric points
isoelectric focusing gel
mixtures of proteins will be separated in the
first dimension by isoelectric focusing, and then the gel will be treated with SDS and
run in the second dimension using SDS-PAGE
two-dimensional gel electrophoresis
If you have a complex mixture of proteins, you can sometimes identify what proteins are in
that mixture using
mass sepctrometry
to determine the structure of large protein complexes, integral membrane proteins and dynamic proteins. This
technique involves rapidly freezing the proteins and exposing them to a beam of
electrons, which projects the images of the molecules onto a detector.
Cryoelectron microscopy
